The studies proposed in this application seek to gain further understanding of fundamental mechanisms of therapeutic and toxic actions of cardiac glycosides. The first objective is to examine in detail the hypothesis that the positive inotropic effects of cardiac glycosides are causally related to inhibition of the monovalent cation transport enzyme complex, NaK-ATPase. Three approaches to this problem are proposed, including: 1) experiments to define changes in myocardial intracellular sodium and calcium concentration using a serial biopsy technique in open-chest dogs; 2) studies of cultured heart cells to define rigorously relationships between digitalis-induced inotropy and altered monovalent cation active transport; and 3) high-resolution 23Na-NMR spectroscopy of living myocardial tissue to determine the time course of changes in intracellular sodium signals, and to relate changes observed to inotropic and toxic effects of cardiac glycosides. Related studies will test the hypothesis that the known sensitivity of subendocardial Purkinje fibers to toxic electrophysiologic effects of cardiac glycosides is due to greater sensitivity to monovalent cation transport inhibition compared to myocardial tissue. A second major objective is to detect and characterize by 31P-NMP spectroscopy the phosphoenzyme intermediate(s) of the putative cardiac glycoside receptor, NaK-ATPase. The hypotheses will be tested that two forms exist for the phosphoenzyme intermediate in the turnover of NaK-ATPase, that these can be resolved by high field pulsed Fourier transform 31P-NMR spectroscopy, and that new information on the interaction of cardiac glycosides with NaK-ATPase can be obtained by this approach.